Pomegranate sarcotesta lectin (PgTeL) inhibits planktonic growth and disrupts biofilm formed by Cryptococcus neoformans.

AIMS: We investigated the putative fungistatic and fungicidal activities of pomegranate sarcotesta lectin (PgTeL) against Cryptococcus neoformans B3501 (serotype D), specifically the ability of PgTeL to inhibit yeast capsule and biofilm formation in this strain. METHODS AND RESULTS: PgTeL showed a minimum inhibitory concentration of 172.0 µg ml-1, at which it did not exhibit a fungicidal effect. PgTeL concentrations of 4.0-256.0 µg ml-1 reduced biofilm biomass by 31.0%-64.0%. Furthermore, 32.0-256.0 µg ml-1 PgTeL decreased the metabolic activity of the biofilm by 32.0%-93.0%. Scanning electron microscopy images clearly revealed disruption of the biofilm matrix. Moreover, PgTeL disrupted preformed biofilms. At concentrations of 8.0-256.0 µg ml-1, PgTeL reduced metabolic activity in C. neoformans by 36.0%-92.0%. However, PgTeL did not inhibit the ability of B3501 cells to form capsules under stress conditions. CONCLUSIONS: PgTeL inhibited biofilm formation and disrupted preformed biofilms, demonstrating its potential for use as an anticryptococcal agent.

Evaluation of Antioxidant, Antibacterial and Enhancement of Antibiotic Action by Punica granatum Leaves Crude Extract and Enriched Fraction against Multidrug-Resistant Bacteria.

The aim of this study was to evaluate the phytochemical composition, antioxidant, and antimicrobial potential of crude extract and fractions of Punica granatum leaves. The extract was produced by turbo extraction, after which hexanic, ethyl acetate, and aqueous fractions were obtained by partitioning. The chemical analyses were performed by thin layer chromatography and high-performance liquid chromatography, and the antioxidant activities were assayed by DPPH. and ABTS.+ . Minimal inhibitory and bactericidal concentrations (MIC/MBC) were applied to twenty-two bacteria. Most strains susceptible to extract/fractions and resistant to antibiotics were selected, and ampicillin, azithromycin, ciprofloxacin, and gentamicin were associated with the ethyl acetate fraction (EAF) against multidrug-resistant strains in modulatory and checkboard models. The data from chromatographic analyses showed flavonoids and tannins in the extract, as well as the enrichment of EAF in phenols, mainly flavonoids. The flavonoids were connected to the electron transfer activity demonstrated in the DPPH. and ABTS.+ assays. Gram-positive strains are more susceptible to EAF. The subinhibitory concentrations of P. granatum enhanced the antimicrobial activity of the agents and reduced the EAF individual MIC, and the combination of EAF and antibiotics demonstrated a synergistic effect. These results present a promising approach for developing a therapy in which antioxidant extracts and fractions can be used in combination with antibiotics.

Antimicrobial potential of Alpinia purpurata lectin (ApuL): Growth inhibitory action, synergistic effects in combination with antibiotics, and antibiofilm activity.

The Alpinia purpurata inflorescence contains a lectin (ApuL), which has immunomodulatory activities on human cells. In the present work, it was evaluated the antibacterial and antifungal effects of ApuL against human pathogens. ApuL showed bacteriostatic activity against non-resistant (UFPEDA-02) and an oxacillin-resistant isolate (UFPEDA-672) of Staphylococcus aureus with minimal inhibitory concentrations (MIC50) of 50 and 400 µg/mL, respectively. In addition, it showed bactericidal effect on the non-resistant isolate (minimal bactericidal concentration: 200 µg/mL). For Candida albicans and Candida parapsilosis, ApuL showed fungistatic effect (MIC50: 200 and 400 µg/mL, respectively). The lectin was able to impair the viability of the microorganism cells, as indicated by propidium iodide (PI) staining. Analysis of growth curves, protein leakage, and ultrastructural changes supported that ApuL acts through distinct mechanisms on S. aureus isolates. Ultrastructural analysis of ApuL-treated Candida cells revealed malformations with elongations and bulges. ApuL-oxacillin combination showed synergistic effect on the oxacillin-resistant isolates UFPEDA-670 and 671, which were not sensitive to lectin alone. Synergism was also detected for ApuL-ceftazidime against a multidrug-resistant isolate of Pseudomonas aeruginosa. Synergistic action of ApuL-fluconazole was detected for C. parapsilosis, which was insensitive to the drug alone. Biofilm formation by S. aureus non-resistant isolate and C. albicans was remarkably inhibited by ApuL at sub-inhibitory concentrations. In conclusion, ApuL showed differential effects on non-resistant and resistant bacterial isolates, was active against Candida species, and showed synergistic action in combination with antibiotics.

Degradation of textile dyes Remazol Yellow Gold and reactive Turquoise: optimization, toxicity and modeling by artificial neural networks.

In this work, the degradation of Remazol Yellow Gold RNL-150% and Reactive Turquoise Q-G125 were investigated using AOP: photolysis, UV/H2O2, Fenton and photo-Fenton. It was found that the photo-Fenton process employing sunlight radiation was the most efficient, obtaining percentages of degradation above 87%. The ideal conditions for the degradation of the dyes were determined from a factorial design 23 and study of the [H2O2] ([H2O2] equal to 100 mg·L-1); [Fe] equal to 1 mg·L-1 and pH between 3 and 4. In the kinetic study, a degradation of more than 97% was obtained after 150 min for the chromophoric groups and 91% for the aromatic compounds. The experimental data obtained presented a good fit to the nonlinear kinetic model. The model of artificial neural networks multilayer perceptron (MLP) (4-11-5) using the software Statistica 8.0 enabled the modeling of the degradation process and showed a better prediction of the data. The toxicity to the seeds of Lactuca sativa and the bacteria Escherichia coli and Salmonella enteritidis allowed to evaluate the effectiveness of the process. The results of this study suggest that the use of photo-Fenton process with sunlight radiation is an effective way to degrade the dyes under study.

Application of the advanced oxidative process on the degradation of the green leaf and purple açaí food dyes with kinetic monitoring and artificial neural network modelling.

The study evaluated the advanced oxidative processes concerning the degradation of green leaf and purple açaí dyes, as well as the prediction of data through artificial neural networks (ANNs). It was verified that percentage of degradation on the wavelengths (λ) of 215, 248, 523 and 627 nm was 5.95, 49.99, 98.17 and 95.99%, respectively, when UV/H2O2 action and UV-C radiation was applied. A non-linear kinetic model proposed by Chan and Chu presented a good fit to the experimental data, reaching an R2 value between 0.978 and 0.999, for the studied λ. Within the ANN simulations through Statistica 6.0, the multilayer perceptron (MLP) (3-9-4) presented a better fit to the experimental data. However, higher values of R² were obtained when utilizing the sklearn package with Python language and an MLP (4-5-4) model. Assays with Staphylococcus aureus and Staphylococcus pyogenes bacteria isolates were performed and it was verified that after employing the UV/H2O2 process, there was a decrease in the toxicity of the solution of dyes. In evaluating S. aureus toxicity, normal growth was observed. However, for S. pyogenes bacteria, it was found that when using the UV/H2O2 process, toxicity was evidenced at post-treatment solution concentrations of 100, 70 and 50%.

A chitin-binding lectin from Moringa oleifera seeds (WSMoL) impairs the digestive physiology of the Mediterranean flour larvae, Anagasta kuehniella.

Biotechnological techniques allow the investigation of alternatives to outdated chemical insecticides for crop protection; some investigations have focused on the identification of molecules tailored from nature for this purpose. We, herein, describe the negative effects of water-soluble lectin from Moringa oleifera seeds (WSMoL) on Anagasta kuehniella development. The chitin-binding lectin, WSMoL, impaired the larval weight gain by 50% and affected the activity of the pest's major digestive enzymes. The commitment of the digestive process became evident after controlled digestion studies, where the capacity of protein digestion was compromised by >90%. Upon acute exposure, the lectin was not resistant to digestion; however, chronic ingestion of WSMoL was able to reverse this feature. Thus, we show that resistance to digestion may not be a prerequisite for a lectin's ability to exert negative effects on larval physiology. The mechanism of action of WSMoL involves binding to chitin with possible disruption to the peritrophic membrane, causing disorder between the endo- and ectoperitrophic spaces. Additionally, results suggest that WSMoL may trigger apoptosis in gut cells, leading to the lower enzymatic activity observed in WSMoL-fed larvae. Although assays employing an artificial diet did not demonstrate effects of WSMoL on A. kuehniella mortality, this lectin may hold potential for exerting insecticide effects on other pest insects, as well for use in other experimental approaches, such as WSMoL-expressing plants. Moreover, the use of WSMoL with other biotechnological tools, such as 'pyramid' crops, may represent a strategy for delaying the evolution of pest resistance to transgenic crops, since its multiple site targets could act in synergism with other insecticide compounds.

Trypsin inhibitor from Moringa oleifera flowers interferes with survival and development of Aedes aegypti larvae and kills bacteria inhabitant of larvae midgut.

Moringa oleifera flower extract, with trypsin inhibitor activity, is a larvicidal agent on Aedes aegypti. This work reports the isolation of trypsin inhibitor (M. oleifera flower trypsin inhibitor (MoFTI)) and its effect on A. aegypti egg hatching, viability of newly hatched larvae, survival of pupae, and growth of inhabitant bacteria from midgut of fourth-instar larvae (L4). MoFTI (K i, 2.4 µM), isolated by affinity chromatography on trypsin-agarose column, was an 18.2 kDa polypeptide on sodium dodecyl sulphate-polyacrylamide gel electrophoresis. Flower extract (at concentrations of 8.5-17.0 mg/mL) reduced egg hatchability while MoFTI (0.05-0.5 mg/mL) did not affect the hatching rate. Mortality of newly hatched larvae ranged from 3.5 to 19.1 % in the presence of the extract (4.0-17.0 mg/mL) and was also promoted by MoFTI (LC50, 0.3 mg/mL). After 72 h, larvae incubated with extract at 13.0 and 17.0 mg/mL were at stages L2 and L1, respectively, while in control they reached L3 instar. In the presence of MoFTI, at all concentrations tested, the larvae did not pass the first instar. Flower extract and MoFTI did not interfere on pupae survival. The extract and MoFTI inhibited the growth of L4 gut bacteria (minimum inhibitory concentrations of 3.47 and 0.031 mg/mL, respectively) but only the inhibitor showed bactericide effect (minimum bactericidal concentration of 1.0 mg/mL). The findings reported herein indicate that MoFTI constitutes a larvicidal principle from M. oleifera flowers against A. aegypti newly hatched larvae and is an antibacterial agent active against the microbiota from L4 gut.

Chemical and antibacterial analysis of Cinnamomum verum leaves extract and fractions against multidrug resistant bacteria.

The chemical composition, the antioxidant and antimicrobial potential of crude extract from leaves Cinnamomum verum and their enriched fractions was studied. Phytochemical analyses were performed by TLC and HPLC, and the antioxidant capacity was verified by DPPH• and ABTS•+. The Minimal Inhibitory/Bactericidal Concentration was conducted against twenty-two bacteria to select five strains susceptible to extracts/fractions and resistant to the antibiotics tested. Interference of Ethyl Acetate Fraction (EAF) in resistance to synthetic antibiotic was assayed by modulatory and checkerboard model. The chromatographic data showed phenolic compounds in crude extract, as well the flavonoid enrichment in the EAF. The combination of EAF and synthetic antibiotics (ampicillin, azithromycin, ciprofloxacin, or gentamicin) provides a synergistic effect against multidrug resistant strains). The results are useful to obtain multi-targeting in a single therapy solution, which on antioxidants molecules plant-derivatives can act synergistically in antimicrobial combinations, a valuable aid as bacterial resistance modifying compounds.

Phytochemical analysis and evaluation of the antimicrobial and antioxidant activities of extracts and fractions of Hymenaea eriogyne Benth.

The objective of this study was to conduct phytochemical characterization and biological evaluation of Hymenaea eriogyne. Crude extracts and fractions from the bark, leaves and pods, were obtained for phytochemical screening by TLC and HPLC, and evaluation of antibacterial and antioxidant potential. Chromatographic data revealed the presence of several metabolites, notably from the flavonoid class. HPLC analysis confirmed the presence of the flavanonol astilbin (taxifolin 3-O-ramnoside) and other flavonoids derived from aglycone taxifolin. In addition, it was possible to quantify phytochemical markers in the extracts and fractions, which showed an increased content of flavonoid and catechin derivatives in the fraction. Better results of the minimum inhibitory/bactericidal concentrations were obtained with extracts and fractions from bark. In the antioxidant activity using the DPPH method, the enriched bark fraction presented an IC50 of 34.46 µg/mL. These results contribute to the continuity of studies on the chemical and biological composition of the species.

Punica granatum sarcotesta lectin (PgTeL) has antibacterial activity and synergistic effects with antibiotics against ß-lactamase-producing Escherichia coli.

The sarcotesta of Punica granatum fruit contains an antimicrobial lectin called PgTeL. In this work, we evaluated the antibacterial activity of PgTeL against five drug-resistant Escherichia coli isolates able to produce ß-lactamases. Minimum inhibitory (MIC) and bactericidal (MBC) concentrations were determined by broth dilution. Morphometric and viability analyses were performed by flow cytometry, and ultrastructural changes were evaluated by scanning electron microscopy. Potential synergistic effects of PgTeL with antibiotics and anti-biofilm effect were also evaluated. PgTeL showed antibacterial activity against all isolates with MIC and MBC values ranging from 12.5 to 50.0 µg/mL and from 25.0 to 100.0 µg/mL, respectively. For most isolates, PgTeL postponed the growth start by at least ten hours. At the MIC, the lectin caused alterations in size, shape and structure of bacterial cells. The combination PgTeL-ceftazidime showed a synergistic effect for all isolates. Synergy was also detected with ampicillin (one isolate), carbenicillin (one isolate), cefotaxime (one isolate), cephalexin (four isolates) and cefuroxime (three isolates). PgTeL exhibited anti-biofilm activity against all isolates, causing ≥50% inhibition of biofilms at or above 6.25 µg/mL. The antibacterial effect of PgTeL and its synergy with antibiotics indicate that this fruit-derived molecule may have potential for future treatment of multidrug-resistant infections.

Nematicidal activity of a water soluble lectin from seeds of Moringa oleifera.

Gastrointestinal nematodes are the major cause of animal diseases, and the indiscriminate use of synthetic anthelmintic contributes to the development of drug resistance. Natural bioproducts are noteworthy since they have proved to be promising for alternative antiparasitic therapies. This study evaluated the in vitro effect of WSMoL (water soluble Moringa oleifera lectin) on hatching of eggs and on the development of early-stage larvae of gastrointestinal nematodes from naturally infected goats. In addition, the interference of WSMoL on activity of proteases was determined and the affinity of the lectin for glycosylated molecules of these parasites was investigated using fluorescein isothiocyanate (FITC)-labeled WSMoL. WSMoL at 250µgmL-1 interfered on hatching of eggs (40.4% of not hatched eggs; p=0.018), and on larval development (stages L1-L3) (IC50 of 78.22µgmL-1). The activity of secreted proteases showed a significant increase in the presence of WSMoL (307.14U/mg-1031U/mg). FITC-labeled WSMoL recognized embryonic egg content and larval content after hatching, which suggests that WSMoL interact with intestinal glycoconjugate receptors in the embryo, as well as in cuticle of the larvae.

PgTeL, the lectin found in Punica granatum juice, is an antifungal agent against Candida albicans and Candida krusei.

The pomegranate (Punica granatum) sarcotesta contains a chitin-binding lectin (PgTeL) with antibacterial activity against human pathogenic species. In this work, the structural stability of PgTeL was evaluated by fluorimetric analysis and the lectin was evaluated for cytotoxicity to human peripheral blood mononuclear cells (PBMCs) and antifungal activity against Candida albicans and Candida krusei. PgTeL folding was impaired when lectin was incubated at pH≥6.0. On the other hand, the lectin did not undergo unfolding even when heated at 100°C. PgTeL (1, 10, and 100µg/mL) was not cytotoxic to PBMCs. Antifungal activity was detected for C. albicans (MIC: 25µg/mL; MFC: 50µg/mL) and C. krusei (MIC and MFC of 12.5µg/mL). Treatment of yeast cells with PgTeL resulted in decrease of intracellular ATP content even at sub-inhibitory concentrations (½MIC and »MIC) and induced lipid peroxidation. In addition, PgTeL damaged the integrity of fungal cell wall of both species, with more pronounced effects in C. krusei. The lectin showed significant antibiofilm activity on C. albicans at sub-inhibitory concentrations (0.195 and 0.39µg/mL). In conclusion, PgTeL is an anti-Candida agent whose action mechanism involves oxidative stress, energetic collapse, damage to the cell wall and rupture of yeast cells.

CasuL: A new lectin isolated from Calliandra surinamensis leaf pinnulae with cytotoxicity to cancer cells, antimicrobial activity and antibiofilm effect.

This work describes the isolation of a lectin (CasuL) from the leaf pinnulae of Calliandra surinamensis and the evaluation of its cytotoxic, antimicrobial and antibiofilm properties. Proteins from pinnulae extract were precipitated with ammonium sulphate (60% saturation) and submitted to Sephadex G-75 chromatography, which yielded isolated CasuL (purification factor: 113). Native CasuL is an acidic protein (pI 5.82) with a relative molecular mass of 48kDa. This lectin is also an oligomeric protein composed of three subunits and mass spectrometry revealed similarities with a Sorghum bicolor protein. CasuL did not undergo unfolding when heated but changes in conformation and hemagglutinating activity were detected at basic pH. CasuL did not reduce the viability of human peripheral blood mononuclear cells but was toxic to leukemic K562 cells (IC50 67.04±5.78µg/mL) and breast cancer T47D cells (IC50: 58.75±2.5µg/mL). CasuL (6.25-800µg/mL) only showed bacteriostatic effect but was able to reduce biofilm formation by Staphylococcus saprophyticcus and Staphylococcus aureus (non-resistant and oxacillin-resistant isolates). CasuL showed antifungal activity against Candida krusei causing alterations in cell morphology and damage to cell wall. In conclusion, the pinnulae of C. surinamensis leaves contain a thermo-stable lectin with biotechnological potential as cytotoxic, antibiofilm, and antifungal agent.

Evaluation of using aluminum sulfate and water-soluble Moringa oleifera seed lectin to reduce turbidity and toxicity of polluted stream water.

Aluminum salts are used as coagulants in water treatment; however, the exposure to residual aluminum has been associated with human brain lesions. The water-soluble Moringa oleifera lectin (WSMoL), which is extracted with distilled water and isolated by chitin chromatography, has coagulant activity and is able to reduce the concentration of metal ions in aqueous solutions. This study evaluated the potential of using aluminum sulfate and WSMoL to reduce the turbidity and toxicity of water from the Cavouco stream located in Recife, Pernambuco, Brazil. The water sample used (called P1) was collected from the stream source, which was found to be strongly polluted based on physicochemical and water quality analyses, as well as ecotoxicity assays with Artemia salina and seeds of Eruca sativa and Lactuca sativa. The assays combining WSMoL and aluminum sulfate were more efficient than those that used these agents separately. Furthermore, the greatest reduction in turbidity (96.8%) was obtained with the treatment using aluminum sulfate followed by WSMoL, compared to when they were applied simultaneously (91.3%). In addition, aluminum sulfate followed by WSMoL treatment resulted in residual aluminum concentration (0.3 mg/L) that was much lower than that recorded after the treatment using only the salt (35.5 mg/L). The ecotoxicity of P1 was also strongly reduced after the treatments. In summary, the combined use of aluminum sulfate and WSMoL was efficient in promoting a strong reduction of turbidity and ecotoxicity of a polluted water sample, without resulting in a high residual aluminum concentration at the conclusion of the treatment.